Image analytic DNA cytometry--a possibility for assessment of prognosis after resection of ductal pancreatic cancers?]

In this study we analysed DNA-ploidy as a potential prognostic parameter in ductal pancreatic carcinoma. Paraffin embedded histological material, obtained by resection from 34 patients with a ductal pancreatic carcinoma, was selected for analysis. Tumor areas within the paraffin embedded material were identified by HE-stained reference sections. One 50 microns section was dewaxed, rehydrated and mechanically and enzymatically prepared to form a suspension of 10000 cells/ml. One milliliter of the suspension, which contained bare nuclei with small rests of cytoplasma, was centrifuged on glass slides. The fixed nuclei were air-dried and stained by Feulgen SITS technique, which allows for the quantitative measurement of DNA. The DNA analysis was carried out with a computer-controlled single-cell cytophotometry. In contrast to using flow cytometry, only the tumor cells were measured by image-cytometry. Overlapping nuclei, dirt and other artifacts as well as inflammatory cells were efficiently eliminated. With DNA image-cytometry, we could differentiate between hypotriploid, triploid, hypertriploid and tetraploid tumors. The DNA-content provided the strongest influence on prognosis in the multivariate analysis.     

DNA-gehalt der tumorzelle

Hepatocellular carcinoma is a heterogeneous disease with considerable differences in malignant behaviour. Some relevant factors for prognosis are known. In this study we analysed DNA ploidy as a potential prognostic parameter. With DNA image cytometry we were able to differentiate between diploid, hypotriploid, triploid, hypertriploid, tetraploid and aneuploid tumours. The best prognosis was for patients with diploid, hypotriploid and tetraploid tumours with a median survival time of 41 months in contrast to 3 months for patients with triploid, hypertriploid or aneuploid tumours. There was a strong correlation between histomorphological parameters and the DNA content. The DNA content of tumour cells may be of considerable clinical relevance in hepatocellular carcinoma regarding the decision as to whether or not to perform a resection. In patients with prognostically unfavourable parameters adjuvant oncological therapy may improve the prognosis.     

Prognostic significance of tumor ploidy and histomorphological parameters in adenocarcinoma of Barrett's esophagus.

BACKGROUND: Despite recent advances in surgical and multidisciplinary treatment, the prognosis for patients with adenocarcinoma of Barrett's esophagus remains poor. The low prognostic accuracy of even surgical pathologic TNM staging suggests that additional parameters are necessary in determining the prognosis. METHOD: In a retrospective analysis of 50 patients who underwent transhiatal or transthoracic esophageal resection due to adenocarcinoma of Barrett's esophagus, a quantitative DNA analysis using image cytometry was performed in addition to the TNM classification and usual morphological criteria. At the time of DNA analysis the histomorphological parameters and survival time were not known. RESULTS: The main prognostic parameter was the curativity (R classification) of the operation. Considering only patients after R0 resection, a multivariate analysis identified the DNA ploidy, the depth of tumor infiltration and distant metastases of prognostically independent factors. Furthermore, within pT2 and pT3 tumors, which account for 80% of all the tumors, DNA ploidy allows an additional prognostic differentiation which is not possible with pT stage alone. CONCLUSION: Patients with a diploid or tetraploid tumor without distant metastasis and a tumor stage pT1-pT3 should have curative (R0) resection. In case of an aneuploid DNA content or a pT4 tumor resection alone shows no advantage as compared to palliative nonoperative procedures.     

Erste Ergebnisse der bildzytometrischen DNS-Analyse zur Beurteilung der Prognose beim papillaren Schilddrüsenkarzinom

Zusammenfassung Bei 19 Patienten, bei denen wegen eines papillären Schilddrüsenkarzinomes eine Resektion durchgeführt worden war, wurde neben der TNM-Klassifikation und den üblichen morphologischen Beurteilungskriterien zusätzlich eine bildanalytische DNS-Zytometrie durchgeführt. Es fanden sich 13 diploide and 6 aneuploide Tumoren. Bei Patienten mit diploiden Tumoren betrug die rezidivfreie Überlebensrate Bowie die Überlebensrate nach 103 Monaten 84% bzw. 92%, wäh-rend 3 von 6 Patienten mit nichtdiploiden Tumoren in nerhalb von 70 Monaten ein Rezidiv erlitten. Zwischen dem DNS-Gehalt der Tumoren einerseits and dem Tumorstadium, der pT-Einteilung, dem Vorhandensein von Lymphknotenmetastasen and einer tumorbedingten Gefäinfiltration andererseits fand sich kein Zusammenhang.

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First results of image cytometric DNS-analysis as a prognostic parameter to papillary thyroid carcinoma

Summary In this study we analysed DNA-ploidy as a potential prognostic parameter in papillary thyreoid carcinoma. Paraffin embedded histological material, obtained by resection from 19 patients with a papillary thyreoid carcinoma, was selected for analysis. Tumor areas within the paraffin-embedded material were identified by HE-stained reference sections. One 50 pin section was dewaxed, rehydrated and mechanically and enzymatically prepared to form a suspension of 10,000 cells/ml. 1 ml of the suspension, which contained bare nuclei with small rests of cytoplasma, was centrifuged on glass slides. The fixed nuclei were air-dried and stained by Feulgen SITS technique, which allows for the quantitative measurement of DNA. The DNA analysis was carried out with a computer-controlled single-cell cytophotometry. In contrast to using flow cytometry, only the tumor cells were measured by image-cytometry. Overlapping nuclei, dirt and other artifacts as well as inflammatory cells were efficiently eliminated. With DNA image-cytometry, we could differentiate between diploid (n = 13) and aneuploid (n = 6) tumors. Best prognosis with a survival rate of 92% after 103 months had patients with diploid tumors in contrast to patients with aneuploid tumors who did not survive more than 72 months.

     

Flow cytometric analysis in colorectal cancer with hepatic metastases and its relationship to metastatic characteristics and prognosis

Nuclear DNA ploidy studies were performed by flow cytometry on extracted nuclei from 65 heptic metastases from colorectal cancer. In 25 patients, both primary and metastatic lesions were available for analysis. Primary carcinomas were DNA diploid pattern in 48.1%, DNA aneuploid in 51.9%. Of 31 hepatic metastases, 11 (35.5%) metastases showed a DNA diploid pattern, and 25 (64.5%) showed a DNA aneuploid pattern. Ploidy pattern was constant between primary and metastases in 80% of tumors. No significant relationship between metastatic characteristics and DNA ploidy pattern was found. The DNA aneuploid cancers had a relatively poorer prognosis in patients with unresectable hepatic metastasis. In resected hepatic metastases from colorectal cancer, rate of hepatic recurrence with DNA diploid pattern was lower than that with DNA aneuploid pattern. Survival of patients with DNA diploid metastases (71% alive at 5 years) was significantly better than that of patients with DNA aneuploid metastases (21% alive at 5 years) (p less than 0.05). These results demonstrated that flow cytometric DNA ploidy measurements may have prognostic value for patients with hepatic metastases from colorectal cancer.     

Automated image analysis DNA cytometry in testicular cancer

The value of automated DNA cytometry for differentiation of testis cancer was evaluated in 54 seminomas, 13 HCG-positive seminomas, and 48 embryonal carcinomas. Slices of paraffin embedded tissue were enzymatically digested and stained with Feulgen SITS after fixation on glass slides. Automated DNA cytometry was performed with a Modular Image Analysis Computer (MIAC). DNA histogram phenotpye and computed DNA indices were correlated with the different tumor types. The ratio of hypertriploid to hypotriploid increased from HCG-positive seminoma over embryonal carcinoma to seminoma. The following mathematical DNA indices were found to correlate with tumor type: mean ploidy, 2c deviation index, 5c exceeding rate, variation coefficient of the GO/1 fraction and DNA nucleus diameter correlation.     

Prognostic significance of DNA aneuploidy in stage I cutaneous melanoma.

The prognostic significance of DNA aneuploidy was studied restrospectively in 177 Stage I cutaneous melanomas. DNA content was determined by flow cytometry of propidium iodide-stained nuclei recovered from formalin-fixed, paraffin-embedded material. Of 162 evaluable histograms, 124 were diploid, 35 aneuploid, and 3 tetraploid. Aneuploidy strongly correlated with established predictors of unfavorable prognosis, namely, thickness p less than .005, level p less than 0.005, ulceration p less than 0.005, and presence of vertical growth phase p less than 0.02. Overall, aneuploidy was strongly correlated with recurrence (p less than 0.005) and shorter disease-free survival (p less than 0.0001). Aneuploidy was an independent predictor of recurrence for tumors less than 1.5 mm thick (p less than 0.0001) and greater than or equal to 3 mm thick (p = 0.031). For melanomas 1.5-2.9 mm thick, aneuploid tumors had a 27% higher recurrence rate than diploid tumors (63% vs. 36%). This was not statistically significant (p = 0.247). In a multivariate analysis of common predictors stratified by thickness, DNA aneuploidy was the most significant independent parameter (p less than 0.002). DNA content appears to be an important stratification parameter for Stage I cutaneous melanoma.     

Prognostic significance of cellular DNA ploidy level of the breast cancer]

This paper reports that cellular DNA ploidy level of paraffin-embedded histological material from 47 patients with stage II breast cancer was measured by flow cytometry. Meanwhile, the influence of cellular DNA level on disease-free survival (DFS) of postoperative patients for the breast cancer was analyzed. The results showed eighteen (38.3%) of tumors examined were DNA diploid, and the remainder were DNA aneuploid. Patients with diploid cancer had a significantly better disease-free survival compared with the patients with aneuploid cancer (P less than 0.02). Our results suggested that cellular DNA ploidy level is a good prognostic factor and of important significance in predicting prognosis of patients with the breast cancer.     

Flow cytometric analysis of DNA ploidy, cell cycle and cytomorphometry in sarcomatoid renal cell carcinoma.

In 5 patients with sarcomatoid renal cell carcinoma, the nuclear DNA content and size were determined by flow cytometry (FCM), and the prognostic value of DNA ploidy, the percentage of S-phase cells (SPF), and the ratio of modal nuclear size with clinicopathologic behavior was analyzed. Age and clinical stage have been shown to have a strong correlation with prognosis. Older patients with a high stage of cancer had poor outcome with a shorter survival time. In all 60% of the tumors were aneuploid. Tumor invasiveness was related to DNA ploidy. With increasing stage, the overall incidence of aneuploid rises. One alive patient had diploid DNA while 75% of the patients who died of sarcomatoid renal cell carcinomas had aneuploid DNA. Diploid sarcomatoid renal cell carcinomas show significantly lower SPF than aneuploid tumors. There was no significant association between the modal nuclear size and the invasiveness of tumors or survival time. This study suggests that FCM analysis of tumor DNA content and cell cycle could be regarded as an additional prognostic determinant of sarcomatoid renal cell carcinoma.     

Single cell photometry analysis of nuclear DNA in treated undifferentiated prostate cancer and its clinical significance

Based on 67 specimens from 20 conservatively treated undifferentiated prostate cancer patients cell nucleic acid analysis combined with a prospective clinical study was attempted using single cell scanning-cytophotometry. Regardless of therapy type a statistically significant difference in nuclear DNA-content was evident between successfully treated prostate cancers and therapy-resistant cancers. Good prognosis was suggested by a significant decline from aneuploid or polyploid to diploid nuclear-DNA-content with a narrow peak in 2c, especially within the first 12 weeks of treatment. Lack of DNA-content alteration or a DNA-frequency distribution change to the right were correlated with rapid clinical regression. Although all had undifferentiated prostate cancer morphology statistically significant differences in nuclear DNA-content were found. The prognostic value of this remains unknown.     

The prognostic significance of nuclear DNA content in non-small-cell lung carcinoma

The nuclear DNA content of paraffin-embedded specimens of primary non-small-cell lung carcinomas was analysed using flow cytometry in 210 patients (80 squamous cell carcinomas, 99 adenocarcinomas, 19 large cell carcinomas and 15 others). The relationship between nuclear DNA content and prognostic factors was studied using multivariate analysis with Cox's proportional hazard model. 1) The frequency of DNA aneuploidy was 77.3% among 210 patients, and it significantly (p less than 0.05) increased with advanced stage and the presence of lymph node metastasis. 2) The patients with DNA aneuploid tumors had a significantly (p less than 0.001) less favorable prognosis than those with DNA diploid tumors among 179 patients with non-small-cell lung carcinomas. Similar results were demonstrated in 79 patients with stage I carcinomas and in 85 patients who underwent absolute curative resection. 3) Multivariate analysis using Cox's proportional hazard model showed that DNA ploidy was an independent prognostic factor for survival. Especially in the patients with absolute curative resection, DNA ploidy was the most important prognostic factor. In conclusion, flow cytometric nuclear DNA content analysis provided useful biological information, and DNA ploidy was an important and major independent prognostic factor in non-small-cell lung carcinoma.     

Evaluation of DNA ploidy combined with a cytometric proliferation index of imprints from core needle biopsies in prostate cancer.

OBJECTIVE: To evaluate if DNA ploidy analysis with a proliferation index (PI) derived from DNA cytometry of imprints from core needle biopsies predicts disease progression in patients with prostate cancer. METHODS: Touch imprints were done on a consecutive series of core needle biopsies taken by the same urologist from 240 patients with suspected prostate cancer, 137 (46%) of whom were found to have prostate cancer and included in the study. Scattered cells to the right of the image cytometry (ICM) ploidy-establishing peak, the S-phase fraction, and those in the G2M area of the ICM DNA histograms, were counted in percent of the total number of tumor cells, this value being designated the ICM PI. Based on previous results in archival fine needle aspiration material, the following classification was used: DNA group I, diploid tumor with a low PI; DNA group II, diploid tumor with an intermediate PI and tetraploid tumor with a low or intermediate PI, and DNA group III, diploid or tetraploid tumor with high PI and all tumors with an aneuploid pattern. RESULTS: Correlation was found to exist between DNA groups I-III and Gleason score (GS) (p < 0.0001), T-stage (p = 0.006), M-stage (p = 0.009) and disease progression (p < 0.0001). Among the 39 patients who had curative treatment and GS 5-7, the progression-free survival rate was 100% in DNA group I, as compared with only 38% in DNA group II and 55% in DNA group III within the follow-up period (p = 0.008). CONCLUSION: DNA ploidy combined with a PI derived from image cytometry of imprints from core needle biopsies yields additional prognostic information in patients with GS 5-7. Diploid tumors with a low PI (DNA group I) are associated with a low risk of disease progression.     

Transitional cell carcinoma of the renal pelvis and ureter: prognostic relevance of nuclear deoxyribonucleic acid ploidy studied by slide cytometry: an 8-year survival time study.

In 72 patients with urothelial carcinoma of the renal pelvis or ureter the ploidy, deoxyribonucleic acid (DNA) heterogeneity and counts of cell cycle phases in the tumor were analyzed by means of single cell DNA cytophotometry with the intention of finding new prognostic factors in addition to those already known (stage and grade). Followup ranged from 1 to 8 years. The results of the DNA analyses were related to the tumor categories, histopathological grading of the tumors and clinical course. Malignancy grade 1 tumors showed DNA frequency peaks in the diploid range, while tumors assessed as malignancy grade 2 showed heterogeneous DNA distribution patterns. Malignancy grade 3 tumors exhibited 71% aneuploid and 29% tetraploid DNA values. The proliferation rate of the tumor cells was statistically significantly higher in malignancy grades 2 and 3 than in malignancy grade 1. The prognosis for grade 1 tumors is good, whereas it is unfavorable in the case of grade 3 tumors. For these 2 groups (patients with grades 1 and 3 tumors) DNA ploidy affords no additional prognostic information. Grade 2 tumors, on the other hand, are heterogeneous in respect to DNA ploidy although they exhibit the same histomorphological degree of differentiation. These tumors can be subclassified as aneuploid (biologically aggressive) and diploid or tetraploid (biologically less aggressive) tumors. There was also a positive correlation between T category and DNA ploidy. The cell lines were aneuploid in 38% of the patients with stage T1 tumors, 56% with stage T2 tumors and almost 85% with stage T3, N+ tumors. A significant correlation was found between the results of DNA cytophotometry and the clinical course of the disease. Patients with diploid tumor cell nuclei had no metastases and no local tumor progression for up to 8 years, whereas patients with aneuploid tumor cell nuclei suffered metastasis and local tumor progression within 24 to 36 months. The patients died of the tumor 36 months after primary diagnosis on the average. The determination of DNA ploidy, tumor heterogeneity and tumor cell proliferation by means of DNA cytophotometry affords valuable clues as to prognosis.     

Prognostic importance of DNA ploidy in medulloblastoma of childhood

The deoxyribonucleic acid (DNA) content of 53 medulloblastomas was analyzed by means of flow cytometry and compared with the clinical and histological findings in the host patients. Analysis of DNA showed that about half of the tumors were diploid and the other half were aneuploid. More diploid tumors were found among patients of a young age, but the difference was without statistical significance. Cellular differentiation of the tumor did not correlate with DNA ploidy. No correlation was found between Chang's T staging system and the DNA ploidy, whereas the M staging correlated with the ploidy; diploid medulloblastomas had a greater tendency to metastasize than aneuploid medulloblastomas (p = 0.0003). Four-year survival was compared with the extent of resection and DNA ploidy. The patients with total resection and aneuploid medulloblastoma had a better prognosis than those with subtotal resection and diploid tumor (p = 0.001). There was only one survivor among eight patients with subtotally resected diploid medulloblastomas, while all of the seven patients with totally resected aneuploid medulloblastomas survived. Comparison of the G0/G1 phase fraction and S phase fraction in the surviving group and the deceased group offered no significant information.     

The relationships among DNA ploidy type determined by laser scanning cytometry, the overexpression of p53 protein and the numerical aberrations of chromosome 7 in bladder cancer

The cellular DNA content of certain malignancies is regarded as a prognostic parameter. The mutant p53 is thought to destabilize centrosome replication, which leads to aberrant mitosis and chromosome instability. We investigated the relationship among DNA ploidy pattern type, numerical aberrations of chromosome 7 and p53 overexpression in 20 transitional cell carcinomas of the urinary bladder. The DNA ploidy pattern type was determined by laser scanning cytometry, while p53 overexpression was investigated immunohistochemically. Using fluorescence in situ hybridization with chromosome-specific probes, the copy number of chromosome 7 was counted by touch preparations of interphase nuclei. The cytometric analysis revealed that the DNA patterns were highly correlated with both the numerical aberrations of chromosome 7 (p = 0.0002) and the overexpression of p53. The incidences of p53 overexpression in DNA aneuploid tumors and DNA diploid ones were 78% and 10%, respectively (p = 0.0017). Our results suggest that the overexpression of abnormal p53 protein induces DNA aneuploidy in bladder cancer.     

DNA ploidy, S-phase fraction and cytomorphometry in relation to survival of human penile cancer.

The prognostic significance of DNA ploidy, DNA index, S-phase fraction (SPF) and median nuclear size was studied in 11 patients with squamous cell carcinoma of the penis. These patients have been followed for a minimum of 7 months after diagnosis. Nuclear DNA content was determined by flow cytometry from paraffin-embedded tissue. Patients with DNA diploid cancer (n = 7, 64%) had a better survival rate than patients with aneuploid cancer, and a small SPF was associated with a favorable outcome. A statistically significant relation between median nuclear size and survival could be demonstrated. Small modal nuclear size associated with poorer prognosis. There was a significant difference in survival between metastatic and nonmetastatic groups of tumors during the follow-up period. This study suggests that flow cytometric determination of nuclear DNA ploidy from paraffin-embedded samples in penile cancer does add an additional prognostic determinant in addition to the clinical staging of tumors.     

Independent clinical and flow cytometric prognostic factors for the survival of patients with stage I gastric cancer

Paraffin-embedded tumor samples from 151 patients with stage I gastric cancer were analyzed by DNA flow cytometry, and 80 patients recieved an infusion of bromodeoxiurudine (BrdU) to determine S-phase fraction. S-phase fractions of tumors were measured by the immunohistochemical method using anti-BrdU monoclonal antibody. Of the 151 patients, 81 (54%), and 70 (46%) showed diploid and aneuploid patterns. There was no significant association between DNA ploidy and wall invasion, histologic type, or lymphatic invasion. Aneuploid tumors were associated with positive-vessel invasion. When the DNA ploidy and clinicopathological parameters were simultaneously entered into the Cox regression model, DNA ploidy and wall invasion emerged as independent prognostic parameters. Aneuploid tumors had significantly higher values of BrdU labeling indices than diploid ones. These results indicate that the determination of DNA ploidy patterns may be an important prognostic factor in patients with stage I gastric cancer, and may be useful in deciding the therapeutic schedule of patients with gastric cancer.     

The clinical usefulness of flow cytometric DNA analysis in prostatic cancer]

Flow cytometry was used to measure the DNA content in archival paraffin-embedded prostatic cancer specimens from 54 patients with known outcomes. The specimens were obtained by transurethral resection of the prostate. DNA ploidy as a predictor of prognosis was compared with histological grade and clinical stage. Although no significant correlation between histological grade or clinical stage and ploidy pattern was demonstrated, an increased percentage of DNA aneuploid tumors was seen in higher histological grade and in advanced clinical stage. The survival rate calculated by Kaplan-Meier analysis showed that DNA ploidy pattern was a more reliable indicator to predict survival probability than histological grade or clinical stage. All patients with a near diploid pattern (11 patients) survived more than 5 years, whereas all those with an aneuploid pattern (21 patients) died within 3.5 years. Of 22 patients with a tetraploid pattern, 15 died of tumor progression within 5 years. The remaining 7 patients with favorable outcome had a relatively lower proliferation index (less than 65) in DNA histogram and none of them suffered from stage D disease. In conclusion, the results from this retrospective study suggest that flow cytometric DNA analysis in prostatic cancer would be useful as a means of providing prognostic information.     

Analysis of flow cytometric nuclear DNA content of the thymoma and its relationship to malignant intensity

Determination was made of the nuclear DNA content of paraffin-embedded specimens of resected thymoma using flow cytometry in 39 patients. Relationships among nuclear DNA content, clinicopathological findings and prognosis were studied. The frequency of DNA aneuploid tumors was 30.8%, 6.7% in stage I (15 patients), 28.6% in stage II (7 patients), 44% in stage III (9 patients) and 62.5% in stage IV (8 patients) according to Masaoka's classification, indicating more in increase with advancing stage and significant (p less than 0.05) more increase invasive thymoma than in noninvasive thymoma. The 5 year and 10 year survival rates of DNA diploid tumors were each 94%, while DNA aneuploid tumors, 75% and 45%. Patients with DNA aneuploid tumors showed less favorable prognosis than those with DNA diploid tumors. Similar results were found in patients with invasive thymoma and in those on whom subtotal or partial resection were performed. The present results indicate DNA aneuploid tumors to have greater malignant intensity than DNA diploid tumors in resected thymoma. Flow cytometric nuclear DNA content analysis provides useful biological data and new indices for evaluating the malignancy of resected thymoma.     

Multiparameter flow cytometric analysis of neoplasms of the central nervous system: correlation of nuclear antigen p105 and DNA content with clinical behavior

Analysis of the DNA content of various solid tumors and hematological malignancies may provide useful prognostic information. To date, however, there has been a striking lack of correlation between DNA content in neoplasms of the central nervous system and clinical behavior. Simultaneous quantitation of DNA content and proliferation-associated nuclear antigen (p105) by flow cytometry was performed on paraffin-embedded tissues representing three major groups of central nervous system neoplasms--1) 21 astrocytic tumors, 2) 13 pituitary tumors, and 3) 19 meningiomas--and the results were correlated with clinical behavior. All 4 well-differentiated gliomas were diploid, while 3 of 9 anaplastic astrocytomas and 1 of 8 glioblastomas had a demonstrable aneuploid peak. Three of 13 pituitary tumors had an identifiable aneuploid peak, while only 2 of 19 meningiomas had an aneuploid DNA content. Cell-cycle analysis of the malignant gliomas revealed a significantly higher proliferative index (PI, %S + G2M) compared with the well-differentiated astrocytomas (P less than 0.05). Within the subgroup of diploid anaplastic astrocytomas, however, extended patient survival appeared to be associated with a higher PI. For diploid pituitary adenomas, the PI was consistently lower in the 3 tumors that recurred than it was in the remaining 8 adenomas. Nuclear antigen quantitation of diploid tumors showed a wide range of p105 expression in G0G1 cells, suggesting that, within each tumor, the cells are heterogeneous with respect to proliferative activity. Aneuploid nuclei of glial tumors showed enhanced expression of p105 relative to diploid cells of the same specimen. In pituitary tumors, the median G2M/G0G1 fluorescence ratio for p105 was significantly higher (P less than 0.05) for the 3 diploid recurrent tumors than for those that did not recur. These data support the assumption that the aggressive clinical course of malignant glial neoplasms may be related to an abnormal DNA stemline and/or an alteration in cell proliferative activity. Cell cycle analysis and measurement of p105 by this technique may provide information useful from both a prognostic standpoint and in directing adjuvant therapy.